菠萝生长素极性运输载体基因AcPINs和AcAUXs的分离与表达分析

乙烯已经被广泛应用于菠萝人工催花,但其分子机制不是很清楚。以菠萝(Ananas comosus L.‘Perola’)茎尖为材料,采用RT-PCR结合RACE方法得到3个生长素极性运输输出载体基因(Ac PIN1、Ac PIN2和Ac PIN3)和2个生长素极性运输输入载体基因(Ac AUX1和Ac AUX2)的c DNA及基因组DNA全长。Ac PIN1、Ac PIN2、Ac PIN3、Ac AUX1和Ac AUX2的c DNA全长分别为2 690、2 388、2 057、2 156和1 580 bp,其开放读码框长度分别为1 854、1 917、1 530、1 479和1 392 bp,分别编码617、638、509、492和463个氨基酸;其基因组DNA全长分别为3 602、3 208、4 204、5 457和2 436 bp,从起始密码子到终止密码子的长度分别为3 244、2 780、3 947、5 264和2 321 bp。氨基酸序列多重比对及系统发育树结果显示Ac PINs和Ac AUXs分别属于植物PINs和AUX/LAXs家族。荧光定量PCR结果表明,菠萝茎尖经200和1 200 mg·L-1乙烯利诱导处理后,Ac PINs的表达上调较多,其中Ac PIN2在处理后的早期(1~2 d)和后期(28~37 d)显著上调,另外两个PIN家族基因Ac PIN1和Ac PIN3在处理后的大部分时间都明显提高。Ac AUX1的上调表达量相对较少,且相对表达量显著提高也主要集中在处理初期(1 d)和后期(28~37 d),而Ac AUX2的上调表达则只在处理后的前期(1、2、9 d)。研究结果表明,乙烯利诱导菠萝成花过程中,存在着生长素极性运输,且生长素极性输出在此过程中的作用可能更重要。

Isolation and Expression Analysis of the Genes Encoding Polar Auxin Transporter AcPINs and AcAUXs in Pineapple

Exogenous ethylene has been widely used to induce pineapple flowering，but the molecular mechanism behind ethephon induction is still unclear. Three genes encoding auxin efflux carriers （designated as AcPIN1，AcPIN2 and AcPIN3），and two genes encoding auxin influx carriers（designated  as AcAUX1 and AcAUX2）were isolated from the pineapple（Ananas comosus L.‘Perola’）shoot apex using RT-PCR and RACE. The full-length cDNA of AcPIN1，AcPIN2，AcPIN3，AcAUX1 and AcAUX2 were 2 690，2 388，2 057，2 156 and 1 580 bp，with the open reading frames of 1 854，1 917，1 530，1 479 and 1 392 bp，which encode a putative protein of 617，638，509，492 and 463 amino acids，respectively. The genomic DNA sequences of AcPIN1，AcPIN2，AcPIN3，AcAUX1 and AcAUX2 were 3 602，3 208，4 204，5 457 and 2 436 bp，with the lengths of 3 244，2 780，3 947，5 264 and 2 321 bp，respectively，from the start codon to the terminator codon. The results of a phylogenetic tree analysis indicated that AcPINs and the AcAUXs belong to plant PINs and AUXs/LAXs groups. Quantitative real-time PCR indicated that the relative expressions of AcPINs were up-regulated much more than AcAUXs after the treatment of 200 and 1 200 mg · L^-1 ethephon. The expression of AcPIN2 was significantly increased at the former period（1–2 d）and the later period（28–37 d）after the treatment. In contrast，the expressions of AcPIN1 and AcPIN3 were significantly up-regulated at most time points after the treatment. On the other hand，the up-regulated expression of AcAUX1 was mainly on the former period（1 d）and later period（28–37 d），while the significantly up-regulated expression of AcAUX2 was only found at the former period（1，2 and 9 d after the treatment）. The results showed that there were polar auxin transport during the flowering of pineapple，and auxin efflux might play a more important role during this process.